Among the 4.2 million adults newly infected with HIV-1, almost 50 % were women. The increased incidence of HIV-1 infections in women, particularly those of childbearing age, underscores the urgent need for effective preventative and therapeutic options that are safe, affordable and culturally accepted. In the absence of an effective preventative vaccine, topical microbicides offer a practical alternative to block HIV-1 transmission at the site of entry, and may provide an useful and cost effective means for preventing spread of virus. Strains of lactobacillus were specifically selected because they can be easily seeded into the vaginal mucosa by oral ingestion, peri-anal transit, or by direct vaginal inoculation. Moreover, colonization with lactobacilli constitutes a normal part of the vaginal microflora, and may protect against the acquisition of sexually transmitted diseases, including HIV. Microbicides based on genetically engineered bacteria offer a significant advantage over other topical applied products in that they can be administered well before sexual contact, and may persist in the vaginal environment for several days following a single administration. The goal of this proposal is to test a panel of HIV-1 cell entry inhibitors, which are engineered for expression and secretion by lactobacilli, as functional microbicides against HIV-1 infection in vitro. Initial studies will focus on membrane fusion inhibitors T-20, T-1249, and 5-Helix. The specific aims of this study are: 1. To genetically engineer lactobacilli to express and secrete HIV-1 entry inhibitors. 2. To determine the antiviral potency of entry inhibitors secreted by Lactobacillus plantarum to inhibit HIV-1 infection. 3. To test naturally occurring human vaginal lactobacilli for effective secretion of entry inhibitors and to optimize their secretion capacity.